Deciphering the Function and Regulation of the B Cell Intrinsic a2,6-Sialylation Network

破译 B 细胞固有的 a2,6-唾液酸化网络的功能和调节

• 批准号: 432173434
• 负责人: Professor Dr. Falk Nimmerjahn
• 金额: --
• 依托单位: Lehrstuhl für Genetik - AG Falk Nimmerjahn
• 依托单位国家: 德国
• 项目类别: Research Units
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/432173434?language=en
• 关键词: Deciphering; Function; Regulation; Cell; Intrinsic

Glycosylation is an essential post-translational modification and plays a critical role for the stability and function of many proteins. In the first funding period the focus of project P3 was to study the impact of a2,6 sialic acid residues on mouse B cell development and function during steady state and activation. To study this in detail, we generated an in vivo model for a B cell specific knockout of ST6Gal1, the enzyme that adds terminal a2,6 sialic acid residues to sugar structures present on antibodies and other cell surface receptors. Using this mouse model allowed to make several interesting observations. Firstly, we were able to demonstrate that IgG sialylation occurs within B cells and not post IgG secretion, as suggested more recently. Moreover, by investigating B cell development in these mice, we show that the number of mature B cell subsets including follicular B cells, marginal zone B cells, germinal centre B cells and plasma blasts/plasma cells in the spleen and B2 cells in the peritoneal cavity was strongly reduced. In line with this defect in development of antibody producing B cell subsets, we noted a severe reduction in serum IgG levels. Surprisingly, a strong IgG subclass specific effect of a deletion of ST6Gal1 became evident during the steady-state. Thus, while serum IgG2c was virtually absent, IgG1, IgG2b and IgG3 levels were only slightly affected, suggesting an impact of a2,6-linked sialic acid residues on IgG subclass switching during the steady state. Upon vaccination with T cell-independent and T cell- dependent antigens, however, a reduced but clearly detectable level of IgG1/IgG2c or IgG2c antigen-specific antibody responses could be elicited, respectively. This may suggest, that co-stimulatory signals via toll-like receptors or other co-stimulatory molecules may be able to overcome the defect in IgG2c switching during the steady state. In the second funding period we would like build on the results of the first funding period and study on a B cell subpopulation and single cell level where and when enzymes determining the level of a2,6-linked protein sialyation are expressed, how different signaling pathways critical for B cell activation and class-switching are affected by a lack of a2,6-linked sialic acid residues and how the phenotypes observed in inbred mouse models can be translated to the human immune system via the generation of humanized mice with a knockdown or knockout of ST6Gal1 in human B cells.

糖基化是一种重要的翻译后修饰，对许多蛋白质的稳定性和功能起着至关重要的作用。在第一个资助期间，P3 项目的重点是研究 a2,6 唾液酸残基对稳态和激活期间小鼠 B 细胞发育和功能的影响。为了详细研究这一点，我们建立了 B 细胞特异性敲除 ST6Gal1 的体内模型，ST6Gal1 是一种将末端 a2,6 唾液酸残基添加到抗体和其他细胞表面受体上存在的糖结构中的酶。使用这种小鼠模型可以进行一些有趣的观察。首先，我们能够证明 IgG 唾液酸化发生在 B 细胞内，而不是像最近提出的那样在 IgG 分泌后发生。此外，通过研究这些小鼠的 B 细胞发育，我们发现成熟 B 细胞亚群（包括脾脏中的滤泡 B 细胞、边缘区 B 细胞、生发中心 B 细胞和浆母细胞/浆细胞）以及腹膜腔中的 B2 细胞的数量大幅减少。与产生抗体的 B 细胞亚群发育中的这一缺陷一致，我们注意到血清 IgG 水平严重降低。令人惊讶的是，ST6Gal1 缺失的强烈 IgG 亚类特异性效应在稳态期间变得明显。因此，虽然血清 IgG2c 实际上不存在，但 IgG1、IgG2b 和 IgG3 水平仅受到轻微影响，表明 α2,6 连接的唾液酸残基对稳态期间 IgG 亚类转换有影响。然而，在用T细胞非依赖性和T细胞依赖性抗原进行疫苗接种后，可以分别引发降低但可清晰检测水平的IgG1/IgG2c或IgG2c抗原特异性抗体应答。这可能表明，通过 Toll 样受体或其他共刺激分子的共刺激信号可能能够克服稳态期间 IgG2c 转换的缺陷。在第二个资助期中，我们希望以第一个资助期的结果为基础，研究 B 细胞亚群和单细胞水平，决定 a2,6 连接蛋白唾液酸化水平的酶表达的位置和时间，对 B 细胞激活和类别转换至关重要的不同信号通路如何受到 a2,6 连接唾液酸残基的影响，以及如何在近交小鼠模型中观察到的表型 通过产生敲低或敲除人类 B 细胞中 ST6Gal1 的人源化小鼠，将其转化为人类免疫系统。